The natural pacemaker of the mammalian heart is the sinoatrial node (SA node) which is located in the high right atrium, and which comprises specialized pacemaker cells that generate electrical impulses characterized by an intrinsic rhythm. The electrical impulse, or pacemaker potential results from the spontaneous depolarization (a bioelectrical process involving the influx and egress of ions which reduces a membrane potential to a less negative value) of the cardiomyocytes within the SA node. This depolarization spreads from the sinus node through the surrounding atrial tissue and then into the atrial-ventricular node (AV node) before proceeding into the ventricular conduction system. Cardiac chronotropic incompetence resulting from cardiac conduction pathway dysfunction results in abnormalities of the cardiac cycle which are commonly referred to as arrhythmias.
It has been demonstrated that xcex2-adrenergic receptors (xcex2AR) regulate cardiac myocyte inotropic and chronotropic responses through a G protein-linked signaling pathway (Holmer, S. R., and Homcy, C. J., Circulation, 84(5):1891-1902 (1991); Inglese, J. et al., J Biol Chem. 268(32):23735-23738 (1993); Lefkowitz, R. J., and Caron, M. G., J Biol Chem. 263(11):4993-6 (1988)). These signaling pathways involve both Gxcex1s-direct and cAMP-mediated interactions with ion channels involved in myocyte depolarization. Stimulation of xcex2AR increases heart rate as well as cardiac inotropic force. Conversely, blockade of xcex2AR decreases heart rate and cardiac contractility. Cardiac chronotropic incompetence is associated with an increased prevalence of morbidity and mortality.
The majority of the causes of chronotropic incompetence require the implantation of an electronic pacemaker, either temporarily or permanently. The dangers of such surgical procedures are well known. Furthermore, electronic pacemaker devices are subject to failure, which necessitates subsequent surgical procedures to replace the defective device. Future treatments for chronotropic incompetence may be based on therapeutics (biological pacemakers) which can specifically enhance the pacemaker potential of endogenous cardiac tissue. Therefore, it is useful to provide novel compositions and alternative methods to alleviate chronotropic incompetence without the necessity of surgical intervention.
The present invention relates to compositions and methods for regulating heart rate comprising localized introduction (e.g., delivery) of one or more exogenous genes to cardiac tissue. Specifically encompassed by the present invention are methods employing gene therapy strategies to provide molecular-mediated or cellular-based biological pacemakers to treat cardiac chronotropic and conduction disorders. The resulting ability to reconstitute the function of defective xcex2-adrenergic signaling cascade in the myocardial tissue localized to the SA node of patients with arrhythmias, cardiac disease, or age-associated myocardial dysfunction offers great hope for the reduction of morbidity and mortality.
As described herein, a murine cDNA chronotropic test system was developed to evaluate the effects of expressing the human xcex22-adrenergic receptor (xcex22AR) in mice under in vitro, ex vivo and finally in vivo conditions. The ability of xcex22AR gene therapy to restore the normal function of endogenous cardiac tissue was further evaluated in a direct porcine cardiac gene therapy system. More specifically, the present invention describes a gene therapy strategy which utilizes localized expression of biological pacemakers to restore the function of the xcex2-adrenergic signaling cascade. The strategy results in improved cardiac performance and is a useful modality to restore the chronotropic and inotropic responsiveness of dysfunctional or senescent mammalian cardiac tissue.
In one embodiment, the biological cardiac pacemaker is a molecularly-mediated pacemaker. The molecularly-mediated pacemaker is an expression construct comprising at least one gene encoding a cellular protein which either upregulates heart rate, alters cardiac rhythm, or encodes a receptor protein or signal transduction molecule which is essential to normal physiologic cardiac conductance. The gene, or genes, are operably linked to expression control sequences. The expression construct comprising the molecularly-mediated pacemaker can mediate either transient or stable expression. For example, the molecularly-mediated pacemakers can be transiently expressed, and can comprise at least one gene selected from the group consisting of a xcex22AR gene, axcex21AR gene, and a Gxcex1s gene. The gene can encode either the endogenous protein or a heterologous protein which is sufficiently homologous to the endogenous protein to possess biological activity in the recipient host cell. In an alternative embodiment, the molecularly-mediated pacemaker can comprise at least one gene selected from the above listed group operably linked to expression control sequences suitable for transient expression under the control of a cardiac tissue specific promoter, which can be either constitutive or inducible. In a further embodiment the cardiac tissue promoter can be specific for atrial tissue.
The invention also pertains to a cellular-based biological cardiac pacemaker utilizing genetically modified cells. A cellular-based cardiac pacemaker can comprise at least one cell transfected or transduced with at least one gene that upregulates heart rate or alters cardiac rhythm, for example a xcex22AR gene a xcex21AR gene or a Gxcex1s gene.
The invention also encompasses methods of regulating in vivo cardiac pacemaking (chronotropic) activity in a mammal by introducing one of the biologic cardiac pacemakers described herein into the SA node region of an endogenous mammalian heart. The mammal for example be a human. The biological pacemaker is introduced into the heart of the mammal, for example, into the right atrium at a site which is localized to a region surrounding the sinoatrial node. The chronotropic method can employ a molecularly-mediated cardiac biological pacemaker comprising at least one gene that upregulates heart rate or alters cardiac rhythm under the control of expression control elements which mediate either transient expression or stable expression, which is either constitutive or inducible.
Cardiac pacemaking activity can also be controlled by a method employing a cellular-based cardiac biological pacemaker comprising at least one myocyte transfected or transduced with at least one gene that upregulates heart rate or alters cardiac rhythm introduced (transplanted or grafted) into the SA node region of the right atria of the recipient host mammal.
The cardiac chronotropy methods described herein can be used for an individual suffering from cardiac conductive tissue incompetence (arrhythmias) indicative of a underlying disorder of cardiac impulse generation, or to treat an older patient experiencing age-related defects in cardiac performance. For example, the method may be useful in clinical conditions characterized by an abnormal sinus rhythm including but not limited to individuals having sick sinus syndrome, sinus bradycardia, or heartblock.
The methods can also be used for permanently regulating cardiac pacemaking activity in a mammal by introducing a stable cellular-based cardiac pacemaker comprising at least one myocyte transfected or transduced with at least one gene that upregulates heart rate or alters cardiac rhythm, or by introducing an molecularly-mediated cardiac pacemaker transcriptionally regulated for stable expression under the control of an inducible promoter.
The invention also encompasses methods of enhancing the basal heart rate of a mammal by delivery into the mammal of a biological pacemaker comprising exogenous genes which upregulate heart rate or alters cardiac chronotropic or inotropic responsiveness. The invention further encompasses methods of enhancing (upregulating) inotropic responsiveness (cardiac function) of cardiac tissue by utilizing one of the biological cardiac pacemakers described herein to upregulate heart rate or cardiac rhythm.
The chronotropic regulatory methods may further employ the in vivo administration of a receptor agonist having a specific cellular affinity for the molecule mediating the chronotropic or inotropic effect. For example, if the activity of the biological pacemaker is based on the expression of xcex22AR, the method could further comprise the systemic or local administration of a cardioselective xcex2-adrenergic agent such as isoproterenol.
Future treatments for chronotropic incompetence may obviate the need for mechanical pacemakers, by employing gene therapy strategies to develop therapeutics (biological pacemakers) which can specifically enhance the pacemaker potential of endogenous cardiac tissue. Therefore, it is useful to provide novel compositions and alternative methods are available to alleviate chronotropic incompetence without the necessity of surgical intervention, and the associated risk of mechanical or electronic failure.